In its most general sense chromatography is the separation of mixtures of chemical substances into their component parts by chromatography adsorption for analytical purposes. Chromatographic adsorption is the preferential adsorption or differential retention of chemical compounds on the basis of molecular size, charge, hydrophobicity or biospecific affinity by an adsorbent material. Liquid chromatography is a form of chromatography using a liquid as the mobile phase and a solid or a liquid on a solid support as the stationary adsorbent phase. A liquid chromatography system typically includes at least the following elements which are connected together in a manner well known to those of ordinary skill in the field: a reservoir, a pump, an injector, a column, a detector, and a recorder. High performance liquid chromatography ("HPLC") utilizes specific modifications in the design and nature of the column, the column material, the stationary phase, the sample/mobile phase injector, and the chromatographic conditions (e.g. pressure, temperature, flow rate) to provide improved separation and resolution for more refined analysis, as will be appreciated by those of ordinary skill in the field.
The mobile phase reservoir is an integral component of any liquid chromatography system, and is particularly critical in an HPLC system. A reservoir in a liquid chromatography system must be capable of performing several functions. It must at least: (1) store the mobile phase; (2) act as the point from which the mobile phase is pumped into the column via the system's pump; (3) act as the site of mixing of the mobile phase to uniformity; (4) act as a reaction chamber in which adjustments to the mobile phase may be performed; (5) act as the site of degassing of the mobile phase; and (6) act as the site of sparging of the mobile phase.
From the chromatographer's point of view, the ability to rapidly mix the mobile phase to uniformity and the ability to utilize as much of the mobile phase as possible without introducing air bubbles into the system are critical features of a liquid chromatography mobile phase reservoir. The known reservoirs for use in a liquid chromatography system are either (1) flat-bottomed reservoirs, (2) reservoirs having a conical base and a single cavity subtending the conical base, or (3) reservoirs having a conical base similar to the Kontes reservoir but terminating in a 1-2 inch diameter flat surface rather than a cavity. In operation a stirrer is placed within the reservoir for mixing the mobile phase to uniformity. In addition, a pump inlet filter is also placed inside the reservoir for removing the mobile phase, and often a sparging device is also inserted to allow for helium sparging. The flat-bottomed reservoirs do not have any cavities, and while they provide good mixing, they require extra care in use in order to avoid interference with the stirrer by the pump inlet filter and the sparging device. Furthermore, these reservoirs must be tilted in order to utilize as much mobile phase as possible, and this procedure is dangerous when volatile or toxic chemicals are used as the mobile phase. Moreover, when the liquid level in these reservoirs falls low enough to expose some of the pump inlet filter to the air or gaseous phase in the reservoir, air bubbles will enter the tubing leading to the pump. Air bubbles cause problems for the pump valves and for the detection downstream from the HPLC column, and are incompatible with an efficient HPLC system.
The known reservoirs having the conical base and a single cavity increase the amount of the mobile phase removed from the reservoir (without resorting to dangerous tilting) compared to the flat-bottomed reservoirs. However, a significant drawback is that mixing is minimal. In these single cavity reservoirs, the cavity holds both a magnetic stirrer and a pump inlet filter. One disadvantage of this single cavity design is that only minimal mixing action for the mobile phase is achieved, even at high stirrer rpms (e.g., 1000 rpm), due to interference from the pump inlet filter and due to the spatial separation of the stirrer from the majority of the mobile phase located in the main chamber of the reservoir. This spatial separation or distance between the majority of the mobile phase and the stirrer is caused by the position of the stirrer at the bottom of the cavity under the inlet filter. For example, a pH adjustment in a typical conical, single cavity reservoir can take 30-60 minutes. Eliminating the cavity but retaining the conical base overcomes the spatial separation problem and improves mixing, but the pump inlet filter must be positioned high up enough on the conical slope to avoid interference with the stirrer. This inlet filter position increases the volume of mobile phase that cannot be pumped out of the reservoir without introducing air bubbles into the system.